During prion disease, aggregated prion protein spreads rapidly throughout the central nervous system, inciting neuronal vacuolation, dystrophic neurites, and ultimately neuronal death. Neuronal expression of the cellular prion protein (PrPC) is required for triggering neuronal death pathways, yet how prions induce neuronal vacuolation and dystrophic neurites is unclear. We have developed a new knock-in mouse model of spongiform encephalopathy due to a single amino acid substitution in the amino terminus of PrPC. Mice develop severe spongiform degeneration and dystrophic neurites, particularly in the hippocampus, and the disease onset is delayed by the co-expression of wild type PrPC. Autophagy-related protein levels are altered in the brain and spinal cord and are similarly altered in infectious prion disease. We aim to decipher the aberrant molecular pathways in this mouse model of spongiform encephalopathy. In Specific Aim 1, we will define the altered cellular and biochemical features associated with spongiform encephalopathy in the transgenic mice during aging. In Specific Aim 2, we will determine the dysregulated endolysosomal signaling pathways that lead to neuronal degeneration, comparing findings with infectious prion disease in mice and sporadic Creutzfeldt- Jakob disease in humans. These studies are the first to target the origin of spongiform degeneration and dystrophic neurites in a knock-in mouse model expressing mutated PrP, and are expected to help define the molecular pathways inciting neuronal death in prion disease.